FAQ: DNA transfection

jetPRIME® is our recommended reagent for day-to-day transfection of adherent cells grown in presence of serum. It is well-suitable for DNA delivery, co-delivery of several plasmids or DNA/siRNA co-transfection, and very gentle on cells since it requires low amounts of reagent and nucleic acid during transfection.

jetPEI® transfection reagent is particularly well suited for automated or manual HTS (High Throughput Screening) with three protocols available: reverse, batch and forward. jetPEI® provides highly reproducible results and the DNA/jetPEI® complexes are stable for up to 4 hours.

For co-transfection of multiple plasmids, we recommend that the total DNA amount per well does not exceed the maximal DNA amount indicated in the jetPRIME® protocol. The ratio to use for each plasmid depends on the size of the plasmid, the plasmid construct and the expected expression level of each plasmid. Each plasmid should represent at least 10% of the total DNA amount per well.

For plasmid DNA/siRNA co-transfection, we recommend jetPRIME® reagent. Please refer to the standard protocol for DNA and siRNA co-transfection available on the jetPRIME page: https://www.polyplus-transfection.com/products/jetprime/

For DNA/ssODN co-transfection (single-stranded oligodeoxynucleotides), the same protocol may be applied.

Some suspension and primary cells, such as lymphocyte cell lines, fibroblasts or neurons, are very difficult to transfect with commonly used DNA transfection reagents.

Polyplus-transfection developed a polymer-based transfection reagent jetMESSENGER®. It was designed for mRNA delivery and showed high efficiency on a wide variety of difficult to transfect cells including lymphoma and blood cell lines, neurons and various primary cells. Compared to DNA, mRNA transfection leads to a high percentage of transfected cells, faster protein expression following transfection, and no risk of genomic insertion in contrast to plasmid or viral vectors.

For optimal transfection efficiency using jetPRIME®, we recommend not exceeding 10 minutes of complexation. Hence, for applications requiring longer incubation times such as High Throughput Screening, we recommend using jetPEI®, as jetPEI®/DNA complexes are stable for up to 4 hours.

There is no size restriction for DNA transfection using jetPRIME®. However, it should be pointed out that for the same amount of DNA, the gene copy number will be lower for large DNA constructs than for smaller plasmids.

jetPRIME® interacts with DNA, forming positively charged complexes which penetrate into the cell through endocytosis. Endosomes release DNA in the cytosol via the proton sponge mechanism (Behr, Chimia 1997). The plasmid mostly reaches the nucleus during mitosis, when nuclear envelope disappears. For this reason, primary cells or cell lines with slow cell division rates are usually hard-to-transfect.

For more information about transfection mechanisms, please consult: https://www.polyplus-transfection.com/technologies/what-is-transfection/